A replication-defective, recombinant Sindbis virus vector was utilized in a novel immunization strategy to induce humoral and cellular responses against hepatitis C virus (HCV). The recombinant vector, pVaXJ-E1E2, expressing the gene for HCV glycoproteins E2 and E1, was constructed by inserting the E1E2 gene into the replicon pVaXJ, a DNA vector derived from Sindbis-like virus XJ-160. The defective replicon particles, XJ-E1E2, were produced by transfecting BHK-21(E+Capsid) cells, the packaging cell lines for the vector from XJ-160 virus, with pVaXJ-E1E2. Both glycoproteins, E2 and E1, were stably expressed, as indicated by immunofluorescence assay (IFA) and Western blotting. Mice were vaccinated using a prime-boost strategy with XJ-E1E2 particles combined with Freund's incomplete adjuvant via intramuscular injection at 0 and 2 weeks. HCV-specific IgG antibody levels and cellular immune responses were evaluated by IFA and IFN-γ ELISPOT, respectively. The results showed that the defective XJ-E1E2 particles in combination with Freund's incomplete adjuvant induced effective humoral and cellular immune responses against HCV glycoprotein E1 or E2, suggesting that a defective Sindbis particle vaccine is capable of eliciting an effective immune response. These findings have important implications for the development of HCV vaccine candidates.